Introduction
Carcinogenicity of Specific Oral Microbes
The Classical Microbial Suspects: Viruses
Emerging Role of Porphyromonas gingivalis and Fusobacterium nucleatum
The Bacteriome Associated with Oral Cancer: Clinical Studies
Variations in Composition of the Bacteriome Associated with Tumors
Study | Sample size (tumor/control)* | Technology used | Case sample | Control sample | Bacterial taxa associated with the tumors | Bacterial functions associated with the tumors |
---|---|---|---|---|---|---|
Nagy et al. 1998 [57•], Hungary | 21/21 (self) | Cultivation; biochemical identification | Surface swabs: tumors | Surface swabs: contagious normal mucosa | Fusobacterium, Porphyromonas, Actinomyces, and Propionibacterium spp. | N/A |
Tateda et al. 2000 [58], Japan | 205/10 (self) OSCC 68 Others 137 | Detection of Streptococcus anginosus by PCR and Southern-blot | Tumor tissue, gingival smears, and oropharyngeal swabs | Fresh contagious normal tissue | Authors concluded S. sanginosus was associated, but conclusion is not supported by the results (lack of proper control) | N/A |
Morita et al. 2003 [59], Japan | OSCC 38/7 Esophageal 18/6 | Detection of S. anginosus by q-PCR | Fresh tumor tissue | Fresh non-cancerous tissue | S. anginosus with esophageal, but not oral, cancer | N/A |
Mager et al. 2005 [60], USA | 45/229 | Checkboard DNA-DNA hybridization | Unstimulated saliva | Unstimulated saliva | Capnocytophaga gingivalis, Prevotella melaninogenica, Streptococcus mitis | N/A |
Sasaki et al. 2005 [61] Japan | 42/7 | Detection of S. anginosus by PCR | Fresh tumor tissue, dental plaque, and saliva | Fresh tissue: leukoplakia, lymphoma, and rhabdomyosarcoma | Authors concluded S. anginosus was associated, but conclusion is not supported by the results (lack of proper control) | N/A |
Hooper et al. 2006 [62•], UK | 20/12 (self) | Cultivation; isolates identified with 16S rRNA gene sequencing (Sanger) | Fresh tumor tissue: deep and superficial | Fresh contagious normal tissue | Micrococcus luteus, Prevotella melaninogenica, Exiguo-bacterium oxidotolerans, Fusobacterium naviforme, Staphylococcus aureus, Veillonella parvula | N/A |
Hooper et al. 2007 [63], UK | 10/10 (self) | 16S rRNA clone sequencing (Sanger) | Fresh tumor tissue | Fresh contagious normal tissue | Ralstonia insidiosa, Fusobacterium naviforme, Peptostreptococcus micros, Clavibacter michiganensis subsp. tessellarius, Capocytophaga sp. oral strain S3, Prevotella sp. oral clone BE073 | N/A |
Katz et al. 2011 [64], USA | 10/5 | Immunohistochemical staining | FFPE gingival carcinoma tissue | FFPE normal gingival tissue |
Porphyromonas gingivalis
| N/A |
Pushalkar et al. 2011 [65], USA | 3/2 | 16S rRNA (V4–5) amplicon sequencing (454); DGGE | Stimulated saliva | Stimulated saliva | Genera Streptococcus, Rothia, Gemella, Peptostreptococcus, Lactobacillus, Micromonas, and Porphyromonas | N/A |
Pushalkar et al. 2012 [66], USA | 10/10 (self) | 16S rRNA clone sequencing (Sanger); DGGE | Fresh tumor tissue | Fresh contagious normal tissue | Parvimonas sp. oral taxon 110, Eubacterium infirmum, Eubacterium brachy, Gemella haemolysans, Gemella morbillorum, Gemella sanguinis, Johnsonella ignava, Peptostreptococcus stomatis, Streptococcus gordonii, and Streptococcus parasanguinis I | N/A |
Schmidt et al. 2014 [67], USA | 16/8/6 (OSCC/“pre-cancer”/ healthy) | 16S rRNA (V4) amplicon sequencing (454 and Miseq) | Surface swabs: tumors and “pre-cancer” lesions | Surface swabs: contralateral normal; normal mucosa from healthy subjects | Genera Fusobacterium and Prevotella (Streptococcus and Rothia showed inverse association) | N/A |
Al-Hebshi et al. 2015 [68], Yemen | 3/0 | 16S rRNA (V1–3) amplicon sequencing (454) | Fresh tumor tissue | None |
Bacteroides fragilis?
| N/A |
Guerrero-Preston et al. 2016 [69], USA | 17/25 OSCC 6 Oropharyngeal 11 | 16S rRNA (V3-V5) amplicon sequencing (454) | Oral rinse | Oral rinse | Genera Streptococcus, Lactobacillus, Staphylococcus, and Parvimonas—also Fusobacterium by G-test (Haemophilus and Neisseria among others showed inverse association) | N/A |
Hu et al. 2016 [70], China | 16/10/19 (OSCC/“pre-cancer”/healthy) | 16S rRNA amplicon sequencing (Miseq) | Unstimulated saliva | Unstimulated saliva | Genera Bacillus (Streptococcus and Abiotrophia showed inverse association) | N/A |
Al-Hebshi et al. 2017 [71•], Yemen/Saudi Arabia | 20/20 | 16S rRNA (V1-V3) amplicon sequencing (Miseq) | Fresh tumor tissue | Deep epithelium swabs | Fusobacterium nucleatum, Pseudomonas aeruginosa, Campylobacter oral taxon 44, Haemophilus influenza, Campylobacter showae, and Parvimonas micra (Streptococcus mitis, Rothia mucilaginosa, and Haemophilus parainfluenzae among others showed inverse association) | As predicted with PICRUSt: bacterial mobility, flagellar assembly, bacterial chemotaxis, and LPS biosynthesis (DNA repair, glycolysis/gluconeogenesis and biosynthesis of amino acids were enriched in health) |
Amer et al. 2017 [72], Ireland | 36/32 (Leukoplakia/healthy) No OSCC | 16S rRNA (V1–V2) amplicon sequencing (Miseq) | Surface swabs: leukoplakia patients | Surface swabs: contralateral normal; normal mucosa from healthy subjects | Genera Fusobacterium, Leptotrichia, and Campylobacter; Rothia mucilaginosa | N/A |
Börnigen et al. 2017 [73], USA | 112/242 OSCC 43 Oropharyngeal 64 Unknown 5 | 16S rRNA (V4) amplicon sequencing (Miseq) | Oral rinse | Oral rinse | Genus Dialister only (Streptococcus and Rothia showed inverse association) | Results ambiguous: discrepancies between the figure and text |
Guerrero-Preston et al. 2017 [74], USA | As above + 154 subjects from the HMP | Same dataset above re-analyzed, along with HMP data, with Resphera Insight | Oral rinse | Oral rinse | Fusobacterium nucleatum, Streptococcus mutans, Parvimonas micra, and Lactobacillus spp. | N/A |
Lee et al. 2017 [75], Taiwan | 125/124/127 (OSCC/“pre-cancer”/healthy) | 16S rRNA (V4) amplicon sequencing (Miseq) | Unstimulated saliva | Unstimulated saliva | Genera Bacillus, Enterococcus, Parvimonas, Peptostreptococcs, and Slackia | N/A |
Mukherjee et al. 2017 [76], USA | 39/39 (self) All mobile tongue cancer | 16S rRNA (V4) amplicon sequencing (Ion Torrent) | Fresh-frozen tumor tissue | Fresh-frozen adjacent normal tissue | Genera Streptococcus, Rothia, Actinomyces, Enterococcus, and Microococcus (Fusobacterium, Prevotella, Parvimonas, Campylobacter, and Porphyromonas showed inverse association) | N/A |
Shin et al. 2017 [77], USA | 34/24 (self) OSCC 4 Others 30 | 16S rRNA (V4) amplicon sequencing (Ion Torrent) | Fresh-frozen tumor tissue: primary tumor or/and metastatic | Fresh-frozen adjacent normal tissue | No significant difference for OSCC analyzed separately. When all tumors pooled: an increase in Fusobacterium and Parvimonas; a decrease in Streptococcus, Veillonella, Actinomyces, and Rothia | N/A |
Wang et al. 2017 [78], USA | 121/121 (self) OSCC 59 Others 62 | 16S rRNA clone sequencing (Sanger) | Fresh-frozen tumor tissue | Fresh-frozen adjacent normal tissue | Genus Parvimonas only (Actinomyces was inversely associated) | N/A |
Zhao et al. 2017 [79], China | 40/40 (self) | 16S rRNA (V4–V5) amplicon sequencing (Miseq) | Surface swabs: tumors | Surface swabs: self- matched normal mucosa | Genera Fusobacterium, Capnocytophaga, Alloprevotella, Treponema, Campylobacter, Parvimonas, and Dialister (Streptococcus, Veillonenlla, Lautropia, and Rothia showed inverse association) | As predicted with PICRUSt: Translation, metabolism of cofactors and vitamins; metabolism of terpenoids and polyketides; replication and repair |
Hayes et al. 2018 [80•], USA | 129/254 OSCC 41 Oropharyngeal 30 Larynx 58 Nested case-control study in two larger cohort studiesa | 16S rRNA (V3–V4) amplicon sequencing (454) | Oral mouthwash samples (collected at baseline) | Oral mouthwash sample (collected at baseline) | Pooling all cases and controls: genera Kingella and Corynebacterium and species Prevotella nanceiensis, Capnocytophaga leadbetteri and Selenomonas sputigena were associated with decreased risk of HNSCC (Parvimonas micra and Neisseria sicca were associated with reduced risk of OSCC) | N/A |
Lim et al. 2018 [81], Australia | 52/21 OSCC 15 Oropharyngeal 37 Healthy subdivided into low and high risk | 16S rRNA (V6–8) amplicon sequencing (Miseq) | Oral rinse | Oral rinse | Pooling all cases: genus Oribacterium only (Rothia, Haemophilus, Corynebacterium, Paludibacter, Porphyromonas, and Capnocytophaga were inversely associated) | N/A |
Perera et al. 2018 [82], Sri Lanka | 25/27 | 16S rRNA (V1–V3) amplicon sequencing (Miseq) | Fresh tumor tissue | Fresh tissue of fibroepithelial polyp | Campylobacter concisus, Prevotella salivae, Prevotella loeschii, and Fusobacterium oral taxon 204 (Streptococcus mitis, 6 other Streptococcus spp., Rothia spp. and Lautropia mirabilis among others showed inverse association) | As predicted with PICRUSt: LPS biosynthesis, peptidases and carbon fixation in photosynthetic organisms among others (Base excision repair, glycolysis/gluconeogenesis, and biosynthesis of amino acids were enriched in health) |
Yang C et al. 2018 [83•], Taiwan | 197/51 (41 stage 1 + 66 stage 2/3 + 90 stage 4)b | 16S rRNA (V3–V4) amplicon sequencing (Miseq) | Oral rinse | Oral rinse | Fusobacterium periodonticum, Parvimonas micra, Streptococcus constellatus, Haemophilus influenza, and Filifactor alocis (in contrast to decrease of Streptococcus mitis, Haemophilus parainfluenzae, Porphyromonas pasteri, Veillonella parvula)—strength of association with staging | As predicted with PICRUSt (in stage 4 vs. health): cytoskeleton proteins, methane metabolism, carbon fixation in photosynthetic organisms, restriction enzymes, others (amino acid synthesis and metabolism were enriched in health) |
Yang S et al. 2018 [84], Taiwan | 39/0 Cases clustered into 3 groups by mutational signature: MSC1 (n = 9) MSC2(n = 10) MSC3 (n = 11) | 16S rRNA (V4) amplicon sequencing (Miseq) | Unstimulated saliva | None | N/A (no control group to compare with: comparisons were made between the three patient clusters) | N/A (no control group to compare with: comparisons were made between the three patient clusters) |
Yost et al. 2018 [85••], USA | 4/4 | Metatranscriptome (mRNA) sequencing (NextSeq 500) | Surface swabs: tumors | Surface swabs: adjacent normal mucosa from the cancer patient; tumor-matching and buccal sites from the healthy subjects | Genera Fusobacteria, Selenomonas, Capnocytophaga, Dialister, and Johnsonella (genus Bacillus; species Porphyromonas catoniae, Kingella denitricans, Capnocytophaga gingivalis, among others, were associated with healthy, tumor-matching sites) | Iron ion transport; tryptophanase activity; peptidases; superoxide dismutase; capsule biosynthesis; flagellum synthesis and assembly; chemotaxis; hemolysins and adhesins (base excision repair only was enriched in health) |
More Consistent Results Obtained with Functional Analysis
The Mycobiome Associated with Oral Cancer
Study | Sample size (tumor/control)* | Technology used | Case samples | Control samples | Key findings |
---|---|---|---|---|---|
Krogh et al. 1987 [88], Denmark | 12/12 (self) Leukoplakia only | Cultivation for isolation HPLC for measuring nitrosation potential | Surface swabs: lesion surface | Surface swabs: adjacent normal mucosa | Candida strains with high nitrosation potential found in lesions with advanced “precancerous” changes |
Rindum et al. 1994 [89], Denmark | 53/100 Leukoplakia 32 CEOC 21 No OSCC | PAS stain for cytological examination Cultivation and biochemical assays for isolation, identification, and typing | Smears and swabs: lesion surface | Smears and surface swabs: adjacent normal mucosa and normal mucosa (cheek, tongue, and palate) from healthy subjects | Greater detection rates (predominantly C. albicans) in pathological lesions than in normal mucosa Rare types of C. albicans tended to be associated with dysplasia and non-homogenous leukoplakia |
Barrett et al. 1998 [90], UK | 4724 mucosal biopsies: OSCC 424 Dysplasia 597 Other lesions 3703 | PAS stain for detection of fungal hyphae | Mucosal biopsy | N/A | Greater detection rates in moderate and severe epithelial dysplasia, median rhomboid glossitis, squamous papillomas, and progressive dysplasia No association with OSCC |
Nagy et al. 1998 [57], Hungary | 21/21 (self) | Cultivation; biochemical identification | Surface swabs: tumors | Surface swabs: contagious normal mucosa |
C. albicans
|
McCullough et al. 2002 [91], UK | 223 subjects: OSCC and dysplasia combined 103 Other mucosal lesions 120 | PAS stain for detection of fungal hyphae; cultivation for isolation and CFU count | Mucosal biopsy; mouth rinse | N/A | Significantly higher prevalence and counts in subjects with OSCC and dysplasia Counts correlated with severity of dysplasia and presence of OSCC |
Spolidorio et al. 2003 [92], Brazil | 832 mucosal biopsies: OSCC 109 Dysplasia 315 Other lesions 408 | PAS stain for detection of fungal hyphae | Mucosal biopsy | N/A | Greater detection rates in epithelial dysplasia, OSCC, and hyperkeratosis |
Hebbar et al. 2013 [93], India | 50 subjects: OSCC 9 Dysplasia 20 Other lesions 21 | PAS stain for detection of fungal hyphae Cultivation for isolation and CFU count | Mucosal biopsy; mouth rinse | N/A | C. albicans prevalence (in both biopsies and mouth rinse) and counts correlated with severity of dysplasia and OSCC (lowest for mild dysplasia, highest for OSCC) |
Berkovits et al. 2016 [94], Hungary | 20/40 | Cultivation for isolation and CFU count MALDI-TOF-MS for identification | Surface swabs: tumor | Surface swabs: adjacent normal mucosa and normal mucosa from healthy subjects | Greater average fungal burden (predominantly Candida) in OSCC vs. healthy and on tumor surface compared to the normal epithelium in the OSCC patients More diverse fungal community in association with OSCC: Candida, Rhodotorula, Saccharomyces, Kloeckera, and others |
Mukherjee et al. 2017 [76], USA | 39/39 (self) All mobile tongue cancer | ITS1 amplicon sequencing (Ion Torrent) | Fresh-frozen tumor tissue | Fresh-frozen adjacent normal tissue | Genera Ophiocordyceps and Guignardia significantly enriched in tumors (Lysurus, Leptosphaeria, Antrodiella, and Boletus more abundant in control samples) |
Perera et al. 2018 [95•], Sri Lanka | 25/27 | ITS2 amplicon sequencing (Miseq) | Fresh tumor tissue | Fresh tissue from fibroepithelial polyp | C. albicans, C. etchellsii, and Hannaella luteola–like species enriched in OSCC (A Hanseniaspora uvarum–like species, Malassezia spp., Aspergillus tamarii, Cladosporium halotolerans, and Alternaria alternata more abundant in control samples) |