16S rRNA amplicon sequencing identifies microbiota associated with oral cancer, human papilloma virus infection and surgical treatment
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Rafael Guerrero-Preston1,2, Filipa Godoy-Vitorino3, Anne Jedlicka4, Arnold Rodríguez-Hilario3, Herminio González3, Jessica Bondy1, Fahcina Lawson1, Oluwasina Folawiyo1, Christina Michailidi1, Amanda Dziedzic4, Rajagowthamee Thangavel5, Tal Hadar1, Maartje G. Noordhuis1,6, William Westra7, Wayne Koch1, David Sidransky1
1Department of Otolaryngology and Head and Neck Surgery, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
2Department of Obstetrics and Gynecology, University of Puerto Rico School of Medicine, San Juan, Puerto Rico
3Natural Sciences Department, Microbial Ecology and Genomics Laboratory, Inter American University of Puerto Rico, Metropolitan Campus, San Juan, Puerto Rico
4Department of Molecular Microbiology and Immunology, Johns Hopkins University School of Public Health, Baltimore, Maryland, USA
5Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, New York, USA
6Department of Otorhinolaryngology-Head and Neck Surgery, University of Groningen, University Medical Center, Groningen, The Netherlands
7Department of Pathology-Surgical Pathology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
Rafael Guerrero-Preston, email: [email protected]
David Sidransky, email: [email protected]
Keywords: microbiome, 16s rRNA, oral cancer, oropharyngeal cancer, human papilloma virus (HPV)
Received: February 23, 2016 Accepted: May 16, 2016 Published: May 30, 2016
Systemic inflammatory events and localized disease, mediated by the microbiome, may be measured in saliva as head and neck squamous cell carcinoma (HNSCC) diagnostic and prognostic biomonitors. We used a 16S rRNA V3-V5 marker gene approach to compare the saliva microbiome in DNA isolated from Oropharyngeal (OPSCC), Oral Cavity Squamous Cell Carcinoma (OCSCC) patients and normal epithelium controls, to characterize the HNSCC saliva microbiota and examine their abundance before and after surgical resection.
The analyses identified a predominance of Firmicutes, Proteobacteria and Bacteroidetes, with less frequent presence of Actinobacteria and Fusobacteria before surgery. At lower taxonomic levels, the most abundant genera were Streptococcus, Prevotella, Haemophilus, Lactobacillus and Veillonella, with lower numbers of Citrobacter and Neisseraceae genus Kingella. HNSCC patients had a significant loss in richness and diversity of microbiota species (p<0.05) compared to the controls. Overall, the Operational Taxonomic Units network shows that the relative abundance of OTU’s within genus Streptococcus, Dialister, and Veillonella can be used to discriminate tumor from control samples (p<0.05). Tumor samples lost Neisseria, Aggregatibacter (Proteobacteria), Haemophillus (Firmicutes) and Leptotrichia (Fusobacteria). Paired taxa within family Enterobacteriaceae, together with genus Oribacterium, distinguish OCSCC samples from OPSCC and normal samples (p<0.05). Similarly, only HPV positive samples have an abundance of genus Gemellaceae and Leuconostoc (p<0.05). Longitudinal analyses of samples taken before and after surgery, revealed a reduction in the alpha diversity measure after surgery, together with an increase of this measure in patients that recurred (p<0.05). These results suggest that microbiota may be used as HNSCC diagnostic and prognostic biomonitors.
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