Skip to main content

Advertisement

SpringerLink
Log in

Characterization of Oral Bacterial Composition of Adult Smokeless Tobacco Users from Healthy Indians Using 16S rDNA Analysis

  • Human Microbiome
  • Published:
Microbial Ecology Aims and scope Submit manuscript

Abstract

The present investigation is aiming to report the oral bacterial composition of smokeless tobacco (SLT) users and to determine the influence of SLT products on the healthy Indian population. With the aid of the V3 hypervariable region of the 16S rRNA gene, a total of 8,080,889 high-quality reads were clustered into 15 phyla and 180 genera in the oral cavity of the SLT users. Comparative analysis revealed a more diverse microbiome where two phyla and sixteen genera were significantly different among the SLT users as compared to the control group (p-value < 0.05). The prevalence of Fusobacteria-, Porphyromonas-, Desulfobulbus-, Enterococcus-, and Parvimonas-like genera among SLT users indicates altered bacterial communities among SLT users. Besides, the depletion of health-compatible bacteria such as Lactobacillus and Haemophilus also suggests poor oral health. Here, the majority of the altered genera belong to Gram-negative anaerobes that have been reported for assisting biofilm formation that leads in the progression of several oral diseases. The PICRUSt analysis further supports the hypothesis where a significant increase in the count of the genes involved in the metabolism of nitrogen, amino acids, and nicotinate/nicotinamide was observed among tobacco chewers. Moreover, this study has a high significance in Indian prospects where the SLT consumers are prevalent but we are deficient in information on their oral microbiome.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

Data Availability

Data can be made available to an individual upon sending an email to the corresponding author. Raw data is also linked with this manuscript.

References

  1. Sinha DN, Gupta PC, Kumar A, Bhartiya D, Agarwal N, Sharma S, Singh H, Parascandola M, Mehrotra R (2018) The poorest of poor suffer the greatest burden from smokeless tobacco use: a study from 140 countries. Nicotine Tob. Res. 20:1529–1532

    Article  PubMed  Google Scholar 

  2. Siddiqi K, Islam Z, Khan Z, Siddiqui F, Mishu M et al (2019) Identification of policy priorities to address the burden of smokeless tobacco in Pakistan: a multi method analysis. Nicotine Tob. Res. 29:ntz163

    Google Scholar 

  3. WHO Factsheet India 2018. Received from regional office of South East Asia.

  4. Rodgman A, Perfetti TA (2009) The chemical components identified in tobacco and tobacco smoke prior to 1954: a chronology of classical chemistry. Beitr. Tabakforsch. Int. 23:277–333

    CAS  Google Scholar 

  5. Vishwakarma A, Verma D (2020) Exploring the microbiome of smokeless tobacco. In: Chaudhary, Raj, Verma, Akhter (eds) Microorganisms for Sustainable Environment and Health. Elsevier, Amsterdam, pp 167–178

    Chapter  Google Scholar 

  6. Han J, Sanad YM, Deck J, Sutherland JB, Li Z, Walters MJ (2016) Bacterial populations associated with smokeless tobacco products. Appl. Environ. Microbiol. 82:6273–6283

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Law AD, Fisher C, Jack A, Moe LA (2016) Tobacco, microbes, and carcinogens: correlation between tobacco cure conditions, tobacco-specific nitrosamine content, and cured leaf microbial community. Microb. Ecol. 72:120–129

    Article  CAS  PubMed  Google Scholar 

  8. Halboub E, Al-Akhali MS, Alamir AH, Homeida HE, Baraniya D et al (2020) Tongue microbiome of smokeless tobacco users. BMC Microbiol. 20:201

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. National cancer institute and centers for disease control and prevention (2014) Smokeless tobacco and public health: a global perspective. U.S. Department of health and human services, Centers for disease control and prevention and National Institutes of Health, National cancer institute. NIH Publication No, Bethesda, MD, pp 14–7983

    Google Scholar 

  10. Miluna S, Rostoka D, Skadioo I, Reinis A, Priedite V, Koka R, Lauva D, Krojea J (2017) The oral microbiome of smokeless tobacco users in Latvia. Proc. Latv. Acad. Sci. 71:33–37

    Google Scholar 

  11. Ganesan SM, Joshi V, Fellows M, Dabdoub SM, Nagaraja HN (2017) A tale of two risks: smoking diabetes and the subgingival microbiome. ISME J 11:2075–2089

    Article  PubMed  PubMed Central  Google Scholar 

  12. Bornigen D, Ren B, Pickard R, Li J, Ozer E, Hartmann EM, Xiao W, Tickle T, Rider J, Gevers D, Franzosa EA (2017) Alterations in oral bacterial communities are associated with risk factors for oral and oropharyngeal cancer. Sci. Rep. 7:17686

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  13. Verma D, Srivastava A, Garg PK, Akhter Y, Dubey AK, Mishra SD, Deo SVS (2020) Taxonomic profiling and functional characterization of the healthy human oral bacterial microbiome from the north Indian urban sub-population. Arch. Microbiol. https://doi.org/10.1007/s00203-020-02084-7

  14. Sievers F, Wilm A, Dineen D, Gibson TJ, Karplus K, Li W, Lopez R, McWilliam H, Remmert M, Söding J, Thompson JD, Higgins DG (2011) Fast, scalable generation of high-quality protein multiple sequence alignments using Clustal Omega. Molecular Sys Biol 7:539

    Article  Google Scholar 

  15. Edgar RC, Haas BJ, Clemente JC, Quince C, Knight R (2011) UCHIME improves sensitivity and speed of chimera detection. Bioinformatics 27:2194–2200

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Edgar R (2013) UPARSE: Highly accurate OTU sequences from microbial amplicon reads. Nat. Methods 10:996–998

    Article  CAS  PubMed  Google Scholar 

  17. Bik EM, Eckburg PB, Gill SR, Nelson KE, Purdom EA (2006) Molecular analysis of the bacterial microbiota in the human stomach. Proc. Natl. Acad. Sci. U. S. A. 103:732–737

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Parks DH, Tyson GW, Hugenholtz P, Beiko RG (2014) STAMP: Statistical analysis of taxonomic and functional profiles. Bioinformatics 30:3123–3124

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Langille MG, Zaneveld J, Caporaso JG (2013) Predictive functional profiling of microbial communities using 16S rRNA marker gene sequences. Nat. Biotechnol. 31:814–821

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Prodan A, Tremaroli V, Brolin H, Zwinderman AH, Nieuwdorp M, Levin E (2020) Comparing bioinformatic pipelines for microbial 16S rRNA amplicon sequencing. PLoS One 15:e0227434

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Bokulich NA, Kaehler BD, Rideout JR, Dillon M, Bolyen E, Knight R, Huttley GA, Gregory Caporaso J (2018) Optimizing taxonomic classification of marker-gene amplicon sequences with QIIME 2’s q2-feature-classifier plugin. Microbiome 6:90

    Article  PubMed  PubMed Central  Google Scholar 

  22. Liu M, Jin J, Pan H, Feng J, Cerniglia CE, Yang M, Chen H (2016) Effect of smokeless tobacco products on human oral bacteria growth and viability. Anaerobe 42:152–161

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Camelo-Castillo AJ, Mira A, Pico A et al (2015) Subgingival microbiota in health compared to periodontitis and the influence of smoking. Front. Microbiol. 6:119

    Article  PubMed  PubMed Central  Google Scholar 

  24. Mason MR, Preshaw PM, Nagaraja HN, Dabdoub SM, Rahman A, Kumar PS (2015) The subgingival microbiome of clinically healthy current and never smokers. ISME J 9:268–272

    Article  PubMed  Google Scholar 

  25. Valles Y, Inman CK, Peters BA (2018) Types of tobacco consumption and the oral microbiome in the United Arab Emirates Healthy Future (UAEHFS) Pilot Study. Sci. Rep. 8:11327

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  26. Verma D, Garg PK, Dubey AK (2018) Insights into the human oral microbiome. Arch. Microbiol. 200:525–540

    Article  CAS  PubMed  Google Scholar 

  27. Jenkinson HF, Lamont RJ (2005) Oral microbial communities in sickness and in health. Trends Microbiol. 13:589–595

    Article  CAS  PubMed  Google Scholar 

  28. Gupta S, Gupta R, Sinha DN, Mehrotra R (2018) Relationship between type of smokeless tobacco and risk of cancer: a systematic review. Indian J. Med. Res. 148:56–76

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Lundmark A, Hu Y, Huss M, Johannsen G, Andersson AF, Yucel-Lindberg T (2019) Identification of salivary microbiota and its association with host inflammatory mediators in periodontitis. Front. Cell, Microbiol 9:216

    Article  CAS  Google Scholar 

  30. Huang C, Shi G (2019) Smoking and microbiome in oral, airway, gut and some systemic diseases. J. Transl. Med. 17:225

    Article  PubMed  PubMed Central  Google Scholar 

  31. Ho D, Ang G, Er C (2018) An unusual presentation of Parvimonas micra infective endocarditis. Cureus 10:e3447

    PubMed  PubMed Central  Google Scholar 

  32. Tindall B, Euzeby J (2006) Proposal of Parvimonas gen. nov. and Quatrionicoccus gen. nov. as replacements for the illegitimate, prokaryotic, generic names Micromonas Murdoch and Shah 2000 and Quadricoccus Maszenan et al 2002, respectively. Int. J. Syst. Evol. Microbiol. 56:2711–2713

    Article  CAS  PubMed  Google Scholar 

  33. Murphy EC, Frick IM (2013) Gram-positive anaerobic cocci – commensals and opportunistic pathogens. FEMS Microbiol. Rev. 37:520–553

    Article  CAS  PubMed  Google Scholar 

  34. Siqueira Jr JF, Rocas IN (2006) Catonella morbi and Granulicatella adiacens: new species in endodontic infections. Oral Surg. Oral Med. Oral Pathol. Oral Radiol. Endod. 102:259–264

    Article  PubMed  Google Scholar 

  35. Carlier JP, Kouas G, Han XY (2007) Moryella indoligenes gen. nov., sp. nov., an anaerobic bacterium isolated from clinical specimens. Int. J. Syst. Evol. Microbiol. 57:725–729

    Article  CAS  PubMed  Google Scholar 

  36. Costalonga M, Herzberg MC (2014) The oral microbiome and the immunobiology of periodontal disease and caries. Immunol. Lett. 162:22–38

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Chen X, Winckler B, Lu M, Cheng H, Yuan Z, Yang Y (2015) Oral microbiota and risk for esophageal squamous cell carcinoma in a high- risk area of China. PLoS One 10:e0143603

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  38. Komiyama EY, Lepesqueur LSS, Yassuda CG, Samaranayake LP, Parahitiyawa NB, Balducci I (2016) Enterococcus species in the oral cavity: prevalence, virulence factors and antimicrobial susceptibility. PLoS One 11:e0163001

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  39. Hung WL, Wade WG, Boden R, Kelly DP, Wood AP (2011) Facultative methylotrophs from the human oral cavity and methylotrophy in strains of Gordonia, Leifsonia, and Microbacterium. Arch. Microbiol. 193:407–417

    Article  CAS  PubMed  Google Scholar 

  40. Tsuzukibashi O, Uchibori S, Kobayashi T, Saito M, Umezawa K, Ohta M, Shinozaki-Kuwahara N (2015) A selective medium for the isolation of Microbacterium species in oral cavities. J. Microbiol. Methods 116:60–65

    Article  PubMed  Google Scholar 

  41. Romanowska I, Kwapisz E, Mitka M, Bielecki S (2010) Isolation and preliminary characterization of a respiratory nitrate reductase from hydrocarbon-degrading bacterium Gordonia alkanivorans S7. J. Ind. Microbiol. Biotechnol. 37:625–629

    Article  CAS  PubMed  Google Scholar 

  42. Zhang D, Li W, Huang X, Wen Q, Liu M (2013) Removal of ammonium in surface water at low temperature by a newly isolated Microbacterium sp. strain SFA13. Bioresour. Technol. 137:147–152

    Article  CAS  PubMed  Google Scholar 

  43. Fisher MT, Bennett CB, Hayes A, Kargalioglu Y, Knox BL, Xu D, Muhammad-Kah R, Gaworski CL (2012) Sources of and technical approaches for the abatement of tobacco specific nitrosamine for mationin moist smokeless tobacco products. Food Chem. Toxicol. 50:942–948

    Article  CAS  PubMed  Google Scholar 

  44. Wei X, Deng X, Cai D, Ji Z, Wang C, Yu J, Li J, Chen S (2014) Decreased tobacco-specific nitrosamines by microbial treatment with Bacillus amyloliquefaciens DA9 during the air-curing process of burley tobacco. J. Agric. Food Chem. 62:12701–12706

    Article  CAS  PubMed  Google Scholar 

  45. Junemann S, Prior K, Szczepanowski R (2012) Bacterial community shift in treated periodontitis patients revealed by Ion Torrent 16S rRNA gene amplicon sequencing. PLoS One 7:e41606

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  46. Chen WP, Chang SH, Tang CY, Liou ML, Tsai SJ, Lin YL (2018) Composition analysis and feature selection of the oral microbiota associated with periodontal disease. Biomed. Res. Int. 2018:3130607

    PubMed  PubMed Central  Google Scholar 

  47. Greabu M, Totan A, Miricescu D, Radulescu R, Virlan J, Calenic B (2015) Hydrogen sulfide, oxidative stress and periodontal diseases: a concise review. Antioxid (Basel) 5:3

    Article  CAS  Google Scholar 

  48. Al-hebshi N, Alharbi F, Mahri M, Chen T (2017) Differences in the bacteriome of smokeless tobacco products with different oral carcinogenicity: compositional and predicted functional analysis. Genes 8:106

    Article  PubMed Central  CAS  Google Scholar 

  49. Hyde ER, Andrade F, Vaksman Z, Parthasarathy K, Jiang H, Parthasarathy DK (2014) Metagenomic analysis of nitrate-reducing bacteria in the oral cavity: implications for nitric oxide homeostasis. PLoS One 9:e88645

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  50. Nasrin S, Chen G, Watson CJW, Lazarus P (2020) Comparison of tobacco-specific nitrosamine levels in smokeless tobacco products: high levels in products from Bangladesh. PLoS One 15:e0233111

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Guerrero-Preston R, Godoy-Vitorino F, Jedlicka A, Rodríguez-Hilario A, Gonzalez H, Bondy J (2016) 16S rRNA amplicon sequencing identifies microbiota associated with oral cancer, human papilloma virus infection and surgical treatment. Oncotarget 7:51320–51334

    Article  PubMed  PubMed Central  Google Scholar 

  52. Lim Y, Fukuma N, Totsika M, Kenny L, Morrison M, Punyadeera C (2018) The performance of an oral microbiome biomarker panel in predicting oral cavity and oropharyngeal cancers. Front. Cell. Infect. Microbiol. 8:267

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  53. Segata N, Haake SK, Mannon P (2012) Composition of the adult digestive tract bacterial microbiome based on seven mouth surfaces, tonsils, throat and stool samples. Genome Biol. 13R42:1–18

    Google Scholar 

  54. Bolourian A, Mojtahedi Z (2018) Streptomyces, shared microbiome member of soil and gut, as ‘old friends’ against colon cancer. FEMS Microbiol. Ecol. 1:94

    Google Scholar 

  55. Li Y, Tan X, Zhao X, Xu Z, Dai W, Duan W, Huang S, Zhang E, Liu J, Zhang S, Yin R, Shi X, Lu Z, Pan Y (2020) Composition and function of oral microbiota between gingival squamous cell carcinoma and periodontitis. Oral Oncol. 107:104710

    Article  CAS  PubMed  Google Scholar 

  56. Bhushan B, Yadav AP, Singh SB (2019) Diversity and functional analysis of salivary microflora of Indian Antarctic expeditionaries. J. Oral Microbiol. 11:1581513

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Wu J, Peters BA, Dominianni C, Zhang Y, Pei Z (2016) Cigarette smoking and the oral microbiome in a large study of American adults. ISME J 10:2435–2446

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Code availability

Not applicable

Funding

The investigation is financially supported by SERB, New Delhi (File No. SB/YS/LS-102/2014), and UGC-BSR (F 30.442/2018/BSR).

Author information

Authors and Affiliations

  1. Department of Microbiology, Babasaheb Bhimrao Ambedkar University, Lucknow, India

    Ankita Srivastava & Digvijay Verma

  2. Department of Bio-Statistics and Data Management, ICMR-National Institute of Occupational Health, Ahmedabad, India

    SukhDev Mishra

Authors
  1. Ankita Srivastava
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. SukhDev Mishra
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Digvijay Verma
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Digvijay Verma.

Ethics declarations

Ethics approval

The protocol of the study was approved by the Institutional Ethical Committee, Netaji Subhas University of Technology, New Delhi (NSUT), as well as the All India Institute of Medical Sciences (AIIMS), New Delhi (Ref No. IEC/NP 166/2014/ RP-14/2014).

Consent to participate

A signed consent was taken from all the participants.

Consent for publication

On acceptance of the manuscript, the copyrights will be transferred to the publisher.

Conflict of interest

The authors declare no competing interests.

Supplementary Information

ESM 1

Supplementary fig. 1. The PICRUSt analysis at tier I, II reveals significant differences among the genes involved in human diseases, and cellular processes (a), the metabolism of amino acids, xenobiotics (b). Supplementary fig. 2. Beta diversity analysis among SLT users (HTC) and non-users (HI) using weighted Unifrac (quantitative) phylogenetic metric pairing method showing less difference at the genera levels. (PDF 356 kb)

ESM 2

(DOCX 45 kb)

ESM 3

(XLSX 82 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Srivastava, A., Mishra, S. & Verma, D. Characterization of Oral Bacterial Composition of Adult Smokeless Tobacco Users from Healthy Indians Using 16S rDNA Analysis. Microb Ecol 82, 1061–1073 (2021). https://doi.org/10.1007/s00248-021-01711-0

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00248-021-01711-0

Keywords

Search

Navigation