Abstract 870: 16S rRNA saliva analysis unveils microbiome biomonitors linked to human papilloma virus and oropharyngeal squamous cell carcinoma
Authors: Guerrero-Preston Rafael E. , Michailidi Christina , Jedlicka Anne , Thangavel Rajagowthamee , Hadar Tal , Nordhuis Maartje G. , Westra William , Koch Wayne , Califano Joseph , Sidransky David 
Published: May 2015

Abstract
Microbiota markers in saliva able to distinguish HPV positive from HPV negative Head and Neck Squamous Cell Carcinoma (HNSCC) patients, as well as HNSCC patients from controls, will enable increases in treatment options, reductions in treatment costs and improvements in survival rates. In the present study we compared the microbiota present in saliva DNA from HPV positive and HPV negative oropharyngeal cancer (OPSCC) patients to saliva DNA from uvulopalatopharyngoplasty (UPPP) patients, used as normal oral mucosa controls.
PCR amplification of the 16S rRNA V3-V5 gene region was performed using the 357F/926R primer set. Amplified fragments were multiplexed on the Roche/454 GS Junior sequencing platform. Data were screened for chimeric sequences and contaminant chloroplast DNA after pre-processing. Passing sequences were characterized for diversity and taxonomic composition using QIIME and R before cross-tabulation analyses were performed.
Bacteroidetes, Firmicutes, and Proteobacteria dominated the microbiome in our sample set with less frequent presence of Actinobacteria and Fusobacteria members. At lower taxonomic levels, the most abundant genera observed were Streptococcus, Prevotella and Veillonella. We found statistically significant associations in the alpha-diversity estimators of UPPP and OPSCC patients. Specifically, the microbiota communities present in UPPP are significantly enriched in the observed_species category (P&lt;0.001). The raw number of operational taxonomic units (OTUs) was also significantly higher in UPPP when compared to OPSCC patients (p=0.02). Specifically, a threshold of 80 OTUs was enough to discriminate UPPP from OPSCC patients.
At the phylum level, we did not detect a significant difference in taxa of UPPP and OPSCC patients. Yet at the family level, the taxon Fusobacteriales_Leptotrichiaceae is dramatically enriched in UPPP samples when compared to OPSCC (p=0.02) and is present in very high numbers in all UPPP samples. A threshold of 10 sequences assigned to Fusobacteriales_Leptotrichiaceae discriminated UPPP from OPSCC patients. At the genera level we found that the genus Tannerella is exclusively observed in UPPP patients (p=0.03).
At the class level, Bacteroidetes_Flavobacteria appears significantly enriched in HPV- when compared to HPV+ OPSCC patients (p=0.02). A threshold of 5 sequences assigned to Flavobacteria discriminated HPV+ from HPV- OPSCC patients. At the class level, we also observed enrichment of the taxon Bacteroidetes_Flavobacteria in UPPP relative to OPSCC HPV+ patients (p=0.03). And at the genus level, the low-level presence of Aerococcaceae_Abiotrophia discriminated UPPP from OPSCC HPV+ patients (p=0.02).
Our preliminary results suggest that the microbial diversity and taxonomic composition of the microbiota in saliva may be useful diagnostic and early detection biomonitors for OPSCC.
Citation Format: Rafael E. Guerrero-Preston, Christina Michailidi, Anne Jedlicka, Rajagowthamee Thangavel, Tal Hadar, Maartje G. Nordhuis, William Westra, Wayne Koch, Joseph Califano, David Sidransky. 16S rRNA saliva analysis unveils microbiome biomonitors linked to human papilloma virus and oropharyngeal squamous cell carcinoma. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 870. doi:10.1158/1538-7445.AM2014-870